A novel approach for discovering retrotransposons: characterization of a long terminal repeat element in the spoilage yeast Pichia membranaefaciens and its use in strain identification.

Abstract

A novel PCR-based approach designed to detect retrotransposon long terminal repeat (LTR) elements via their association with tRNA genes was applied to Pichia membranaefaciens, an industrially important food spoilage yeast. A single primer based on tRNA gene sequences was used to amplify DNA fragments from different strains, and an observed fragment size difference among strains was found to correspond to the expected size of an integrated LTR. A 289-bp element was cloned as part of the larger fragment and shown to be present in a high copy number and variable genomic location in all strains examined. Sequence analysis revealed the element to be bounded by nucleotides TG at the 5' end and CA at the 3' end and to exhibit target site duplication and other sequence motifs diagnostic of retrotransposon LTRs. LTR sequence data enabled the development of a rapid identification method which distinguished among different strains. The novel method for LTR isolation and the strain identification system are both likely to prove generally applicable for a wide range of other organisms.